The large majority of older patients with autoimmune Graves' hyperthyroidism and atrial fibrillation (AF) have coexisting activating autoantibodies (AA) to the beta1-adrenergic (B1AR) and M2 muscarinic receptors (M2R). Adrenergic and muscarinic agonists are known to enhance the likelihood for developing AF. HYPOTHESIS: AA in the aging thyrotoxic heart facilitates and/or causes AF. We have demonstrated AABAR) in 94% and AAM2R in 88%; and both autoantibodies in 83% in Graves' hyperthyroidism + AF compared to 10% of Graves' patients in normal sinus rhythm (P<0.001). All patients with AF had at least one of the two AA. These autoantibodies altered electrophysiological activity in pulmonary vein atrial sleeve cells such as predisposes to AF in experimental models; supporting our concept that elevated AABAR and AAM2R as well as thyroid hormone are major risk factors for AF. PROPOSAL: We will use translational and mechanistic studies to examine the electrophysiological interrelationships of thyroid hormone, age and activating autoantibodies as a cause of AF. METHODS: (Spec. Aim A) Expansion of epidemiological studies to identify the prevalence and function of autoantibodies in non-Graves hyperthyroidism and AF; (Spec. Aim Bi) Identify age and thyroid hormone dependent triggering of ectopic action potentials in the presence of target-specific autonomic AA in a canine pulmonary vein atrial sleeve tissue preparation in vitro; (Spec. Aim Bii) Study the development of AF (by ECG telemetry) in young (6 mo) and old (>5yrs) rabbits immunized to produce target-specific AAB1AR and/or AAM2R without and with thyroid hormone. We will then use sophisticated electrophysiological studies incorporating right atrial ganglia stimulation to measure the pulmonary vein atrial sleeve cell threshold for induction of AF. This model will determine the combined effects of AABAR and AAM2R on the atrial substrate for AF. METHODS also include a FRET-based micro-cAMP assay to examine the allosteric effects of AA. This assay will assist in determining whether these AA act as agonists and whether they also serve as partial antagonists to their normally operative orthosteric ligands. Our studies are NOVEL in that they will identify a spectrum of activating autoantibodies in the heart and demonstrate their role in immune-mediated mechanism(s) leading to AF in hyperthyroidism. Our data are relevant not only to hyperthyroidism but will also lead to a better understanding of these mechanisms in other and more common forms of AF which frequently coexist with AA. POTENTIAL IMPACT ON VETERANS HEALTH CARE: AF is an important risk factor for stroke and heart failure in our aging veteran population and leads to increased mortality and morbidity. The present study addresses this issue and may permit identification of future therapeutic options.